PROPOSED PROGRAM The purpose and central theme of this revised program continues to be molecular mechanisms that control thrombus formation and dissolution. The experimental strategies include the use of classical biochemical approaches to purify and characterize specific domains or proteins, the use of hybridoma technology to develop monoclonal antibodies to specific protein domains, DNA technology to express normal and mutated domain/proteins, newly evolving peptide synthetic and analytical chemistry to totally synthesize/analyze domains, and molecular modelling to obtain preliminary insights into the 3- dimensional structure of these domains/proteins, as well as crystallography and x-ray diffraction analysis to determine the actual 3-dimensional structure of domains/proteins. The program consists of 6 projects and 3 core units. In Project 1, Dr. Luskutoff will continue studies on the fibrinolytic system and plasminogen activator inhibitor (PAI-1) binding proteins by determining the structure and function of the somatomedin B domain of vitronectin which functions as the high affinity binding site for PAI-1 and characterizing novel PAI-1 binding proteins. In Projects 2 and 3, Drs. Ginsberg and Ruggeri will define in molecular detail, reactions involving the binding of major adhesive proteins (fibronectin and fibrinogen) to their integrin receptors on cells or in the matrix, and will collaborate to determine the influence of flow on these interactions, In Project 4, Dr. Madison will study and compare structure/function relationships in tissue type and urokinase-like plasminogen activators, focusing on fundamental mechanisms that govern inhibitor specificity and zymogenicity. In Project 5, Dr. Griffin will continue his studies of the protein C pathway, focusing on structure/function relationships in protein C and activated protein C (APC), and on the purification and characterization of a putative novel APC cofactor. In Project 6, Dr. Getzoff will crystallize and determine the 3-dimensional structure of antithrombin III and molecules that interact with it. The synthetic and analytical protein chemistry core (Core A) and the hybridoma and peptide antibody core (Core B) will be used by each of the projects and will provide highly sophisticated technologies and specific monoclonal antibodies to probe domain:domain interactions. Core C will be responsible for overseeing the scientific and administrative aspects of this project. The proposed studies will elucidate structural and functional characteristics of vascular proteins and their mechanisms of interaction to advance current knowledge of thrombosis and facilitate the identification of novel therapeutic approaches.